We have completed the structural map of the malic enzyme gene. This gene is more than 115 kb long and split into 14 exons. We have also determined the sequence of the 914 base pairs of DNA 5' of the coding region of the malic enzyme gene. This region possesses neither TATA nor CCAAT sequence but it is rich in G/C residues. Promoter activity of chimeric genes that contain various parts of the 5' flanking region of the malic enzyme gene placed upstream of the coding sequence of the chloramphenicol acetyl was tested in transient transfection assays using several cell lines. Deletion analyses have revealed that sequences +1 to -41 are sufficient to initiate expression, although inclusion of sequences up to -177 is necessary for maximal promoter activity. The chromatin structure of the malic enzyme gene has been analyzed in different thyroidal states. There are four hypersensitive sites in the 5' flanking region of the gene at positions -50 bp, -170 bp, -310 bp, and at approximately -4.1 Kb. There are five hypersensitive sites spaced approximately equidistantly triiodothyronine increased the proportion of chromatin displaying malic enzyme gene hypersensitivity. We have shown that this might be due to "recruitment" of a population of hepatocytes after triiodothyronine treatment, by localizing malic enzyme mRNA in hepatocytes using in situ hybridization histochemistry.